1. Field of the Invention
The present invention is directed to covalently linked oligonucleotide-cyclopropapyrroloindole conjugates which selectively hybridize with a target nucleic acid (or fragment thereof) of complementary nucleotide sequence, and which after hybridization efficiently cross-link with appropriately positioned adenine bases in the target sequence.
2. Brief Description of the Prior Art
The concept of using oligodeoxyribonucleotides and oligoribonucleotides (collectively ODNs) which sequence specifically bind to a target nucleic acid (or portion thereof) for therapeutic, diagnostic and analytical (sequence determination/gene mapping) purposes is old in the art. The use of ODNs having one or more covalently bonded cross-linking agents, to sequence specifically hybridize with a complementary target nucleic acid (or portion thereof) and therafter cross-link to the single or double stranded target, is also known in the art. Cross-linking may occur as a result of irradiation of the hybrid, for example when a psoralen moiety serves as the cross-linking agent. Published PCT applications WO 93/03736 and WO 94/17092 (assigned to the same assignee as the present application) describe oligonucleotides which include one or more covalently linked electrophilic cross linking agents, and which bind to a complementary sequence of nucleic acid (or portion thereof) followed by cross-linking (alkylation) of a base in the target sequence. The latter concept is related to the pioneering work of B. R. Baker, "Design of Active-Site-Directed Irreversible Enzyme Inhibitors," Wiley, N.Y., (1967), who used what was termed "active-site-directed enzyme inhibitors" in chemotherapeutic applications.
Important criteria for utilizing ODN-crosslinker conjugates for chemotherapeutic applications include selectivity of the ODN to reversibly bind only to the desired target sequence of nucleic acid, and effectiveness of the cross-linking reaction in terms of velocity and specificity of the reaction with the nucleophilic group in the target. In other words, it is desirable for the electrophilic group to be substantially unreactive under the conditions of the expected use of the ODN-crosslinker conjugate with nucleophiles in the reaction medium (including the cell where the therapeutic agent is expected to act) and to undergo the cross-linking reaction relatively rapidly, but only after the hybridization of the ODN portion of the conjugate to the target sequence has already occurred. Similar criteria apply to the use of ODN-electrophilic cross-linker conjugates for diagnostic and analytical applications.
Whereas, certain ODN-cross-linker conjugates known in the art before the present invention justify reasonable expectations of utility in diagnostic, analytical and chemotherapeutic applications, many suffer from shortcomings in terms of the criteria discussed above. For these and other reasons, there is definite room in the art for improvement in the nature of ODN-crosslinker conjugates for therepeutic, diagnostic and analytical use. The present invention provides such improvement.
Another aspect of the pertinent background of the present invention relates to the antitumor antibiotic dextrorotatory (+) CC-1065, the structure of which is shown in Formula 1. ##STR1##